1. Field of the Invention
This invention relates to the production of immune lymphokines and the use of those lymphokines to combat microbial infections.
2. Description of the Prior Art
Despite the efforts of researchers and public health agencies, the incidence of human salmonellosis has increased over the past 20 years. The number of actual reported cases of human Salmonella infection exceeds 40,000 per year. However, the Communicable Disease Center estimates that the true incidence of human Salmonella infections in the U.S. each year may be as high as 2 to 4 million. Animal food products, including swine, remain a significant source of human infection.
In addition to the impact of Salmonella on human health, Salmonella infections in swine cost the United States swine industry more than 100 million dollars annually (Schwartz, 1990, "Salmonellosis in Midwestern Swine", In: Proceedings of the United States Animal Health Assoc., pp. 443-449). In the U.S., salmonellosis caused by S. choleraesuis, the etiologic agent of swine paratyphoid, occurs most frequently. While pigs can be exposed to a broad host range of salmonellae, such as S. typhimurium, from a variety of sources, S. choleraesuis is a host adapted pathogen rarely isolated from non-swine sources (Schwartz, ibid). Thus, natural infection of new animals by S. cholerasuis occurs primarily via horizontal transmission from infected animals which shed the pathogens from their gastrointestinal tract.
Blecha et al. (1983, J. Anim. Sci., 56:396-400) and Wilcock and Schwartz [Salmonellosis, IN: Diseases of Swine, 7th edition, Leman et al. (eds.), Iowa State University Press, Ames, Iowa, 1992, pp. 570-583] have disclosed that weaned pigs have an increased susceptibility to infectious diseases in comparison to mature and suckling swine. This increase in susceptibility to infectious agents post-weaning may be comprised of multiple factors, including loss of maternally derived antibodies, developmental deficiencies of the immune response, and stress-induced susceptibility due to increased glucocorticoids in these pigs [Blecha et al. (ibid); Wilcock and Schwartz (ibid); Blecha et al. (1985, Am. J. Vet. Res., 46:1934-1937); Aurich et al. (1990, J. Reprod. Fert., 89:605-612); Abughali et al. (1994, Blood, 83:1086-1092); and El-Awar and Hahn (1991, J. Leuk. Biol., 49:227-235].
With the trend leaning towards weaning piglets from sows earlier, 8-10 days of age in some cases, the influence of developmental deficiencies of the immune system on increased susceptibility to infectious diseases becomes an even more important concern (Blecha et al., ibid). Developmental deficiencies in immune functions and subsequent susceptibility to infectious diseases have been well documented in neonatal mammalian species. Human, equine and bovine neonates exhibit deficient or impaired neutrophil and T cell functions for the first weeks of life [Coignal et al. (1984, Am. J. Vet. Res., 45:898-901); Hauser et al. (1986, Am. J. Vet. Res., 47:152-153); Hill (1997, Pediatric Research, 22:375-382); Miller (1979, Pediatrics (suppl.), 709-712); Rosenthal and Cairo (1995, Intern. J. Ped. Hem./Onc., 2:477-487); Higuchi et al. (1997, J. Vet. Med. Sci., 59:271-276); Lee and Roth (1992, Comp. Haem. Intern., 2:140-147); Lee and Kehrli (Am. J. Vet. Res., 59:37-43); and Zwahlen et al. (1992, J. Leuk. Biol., 51:264-269)]. Susceptibility to gram negative bacteria has also been well documented in equine, porcine, and bovine neonates [Carter and Martens (1986, Comp. Cont. Educ. Pract. Vet., 8:S256-S270); Drieson et al. (1993, Aust. Vet. J., 70:259-262); and Selim et al. (1995, Vaccine, 13:381-390)]. Young pigs exhibit developmental deficiencies within both the humoral and cellular arms of the immune system. Development of B and T cell compartments in neonatal pigs takes several weeks to become stable and the different classes of immunoglobulins in various sites change with the age of the pig [Bianchi et al. (1992, Vet. Immun. Immunopath., 33:201-222) and McCauley and Hartmann (1984, Res. Vet. Sci., 37:234-241)]. Decreased mitogenic responses of T cells and decreased neutrophil function from young pigs have also been observed [Blecha et al. (1983, ibid); El-Awar and Hahn (ibid); Shi et al. (1994, J. Leuk. Biol., 56:88-94); and Hoskinson et al. (1990, J. Anim. Sci., 68:2471-2478)].
Considering the widespread presence of Salmonella in the environment, it is unlikely that animals can be completely protected from Salmonella exposure. Therefore, researchers have continued to investigate means of increasing resistance to colonization in animals exposed to Salmonella. Studies have focused on the evaluation of vaccines, establishment of protective normal intestinal flora, and the identification of feed additives that will inhibit Salmonella growth and colonization. The role of host immunity against Salmonella colonization is unclear, and it also remains uncertain if stimulation of immune responses will effectively enhance colonization resistance. Experimental vaccines have not proven to be consistently effective.
Our laboratory has previously focused on developmental deficiencies of the immune response of neonatal avian species during the first 4-to-7 days post-hatch and the possibility of augmenting the immune response during the first week post-hatch and at other times of increased susceptibility to disease. Accompanying these deficiencies in immune functions is an increased susceptibility to bacterial infections [Ziprin et al. (1989, Poult. Sci., 68:1637-1642)].
We have demonstrated that the administration of immune lymphokines (ILK) derived from the splenic T cells of Salmonella enteritidis (SE)-immune chickens protects both chickens and turkeys from SE organ invasion at one day-of-age [McGruder et al. (1993, Poult. Sci., 72:2264-2271); Ziprin et al. (1996, Avian Dis., 40:186-192); and Tellez et al. (1993, Avian Dis., 37:1062-1070)]. Further, heterophils isolated from the peripheral blood of day-old chickens and turkeys treated with ILK exhibit increased functional capabilities, showing increased phagocytic and bactericidal activities [Lowry et al. (ibid); and Kogut et al. (1995, J. Leuk. Biol., 57:56-62)]. These early studies utilizing ILK involved the batchwise production of SEILK from hyperimmunized chickens for continued experimental use. More recently, Kogut et al. (U.S. patent application Ser. No. 08/929,074, and U.S. Pat. No. 5,691,200) described the production of immortalized cell lines from avian T lymphocytes and the use of those cell lines to produce the SEILK.